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A Local Action of Neurotrophin-3 Prevents the Death of Proliferating Sensory Neuron Precursor Cells
Wael M ElShamy, Patrik Ernfors Neuron Volume 16, Issue 5, Pages (May 1996) DOI: /S (00)
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Figure 1 Neuronal Numbers, Proliferation, and Cell Death in the L5 Dorsal Root Ganglion (A) Number of neurons in the dorsal root ganglion of NT-3+/− and NT-3−/− mice as percent of age-matched control mice. Note the pronounced reduction in neuronal numbers between E11 and E12 of the NT-3−/− mice (closed bars). A slight reduction in neuronal numbers was detected in NT-3+/− mice (open bars). (B) Cell death in the NT-3+/− and NT-3−/− mice as percent of age-matched control mice detected with the TUNEL method at indicated embryonic stages. Note the pronounced excessive cell death at E11 and almost normal levels at E17. (C) Number of proliferating cells in the NT-3+/− and NT-3−/− mice as percent of age-matched control mice detected with BrdU immunohistochemistry at indicated embryonic stages. Student's t tests: single asterisk, p < 0.05; double asterisk, p < 0.01; triple asterisk, p < Neuron , DOI: ( /S (00) )
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Figure 2 Cell Death in the Dorsal Root Ganglion
(A and B) Histology of the dorsal root ganglion. Photomicrographs of E11 control (A) and NT-3−/− (B) lumbar dorsal root ganglia stained with cresyl violet. Note the reduction in ganglion size already at E11 of NT-3−/− mice. (C, D, G, and H) Cell death in the lumbar dorsal root ganglia. Photomicrographs of E11 (C) and E17 (G) control and E11 (D) and E17 (H) NT-3−/− lumbar dorsal root ganglia stained with the TUNEL method. Note numerous apoptotic cells at E11 in the NT-3−/− ganglia (D; arrows). (E, F, I, and J) Proliferation in the lumbar dorsal root ganglion. BrdU immunohistochemistry of E11 (E) and E17 (I) control, and E11 (F) and E17 (J) NT-3−/− lumbar dorsal root ganglia. Arrows indicate some BrdU+ cells. Note the reduction in the number of stained cells in the E11 NT-3−/− lumbar dorsal root ganglion (F). Scale bar in (A) and (B) is 80 μm and in (C)–(J) is 40 μm. Neuron , DOI: ( /S (00) )
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Figure 3 Dying Dorsal Root Ganglion Cells of the NT-3−/− Mice Incorporate BrdU, but Do Not Express Peripherin Photomicrographs are from E11 NT-3−/− mice. (A) Staining with the TUNEL method and for incorporation of BrdU, marking proliferating cells. TUNEL staining is green and BrdU is red and both stain the nucleus. Note the presence of numerous double stained cells (light yellow) indicated by open arrows and the staining of some cells with only the TUNEL method (arrows). (B) Staining with the TUNEL method and for expression of peripherin, a marker for postmitotic neurons. TUNEL staining is green and localized to the nucleus (arrows), and peripherin is red and localized to the cytoplasm (arrowheads). Only rarely was peripherin present in dying cells. One such cell is indicated by a curved arrow. Scale bar in (A) and (B) are 20 μm. Neuron , DOI: ( /S (00) )
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Figure 4 RNase Protection Assay for the Detection of NT-3 mRNA Expression in Freshly Dissected Dorsal Root Ganglia at Embryonic and Postnatal Stages Yeast RNA was used as a negative control for the protection. Note NT-3 mRNA expression in the dorsal root ganglion at E11 and E17, but not postnatally. A GAPDH probe was used to assess mRNA integrity and to compare loading among the lanes. Neuron , DOI: ( /S (00) )
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Figure 5 A Local Source of NT-3 Prevents the Cell Death of Cultured Early Sensory Neurons (A) The survival of E11 dorsal root ganglion neurons from control and NT-3−/− mice cultured in the presence or absence of recombinant NT-3 (10 ng/ml). The percent survival of neurons in the absence of NT-3 compared with in the presence of NT-3 is shown. Note the pronounced loss of NT-3−/− neurons compared with control neurons. (B) Percent survival of wild-type neurons in the presence or absence of BDNF, NT-3, anti-NT-3 antibodies, or a combination of antibody with BDNF or NT-3. Note the pronounced loss of neurons cultured with anti-NT-3 antibodies compared with cultures maintained in the absence of antibodies (no additions). Abbreviations: Rb-NT-3 Ab, rabbit anti-NT-3 antibody; T-NT-3 Ab, turkey anti-NT-3 antibody. Neuron , DOI: ( /S (00) )
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